Our previous studies indicate that muscle enervation occurs approximately 2-3 months prior to symptom onset in the mouse model of amyotrophic lateral sclerosis (ALS). In our preliminary studies, we find that coincident with this early enervation, ultrastructual changes occur throughout the motoneurons (MNs) from neuromuscular presynaptic terminal to distal dendrites. We propose to identify and characterize the earliest pathological changes that occur and to determine the order in which they occur to distinguish initiating events from secondary events. Our overall hypothesis is that systematic examination and characterization of early events associated with initial denervation will provide insight into disease mechanisms resulting in MN dysfunction. ALS is a most debilitating neurodegenerative disease whose underlying causes and pathophysiology are not understood. As a result, there is no treatment that significantly ameliorates or delays the progression of the disease, and death resulting from respiratory failure occurs within 3-5 years from diagnosis. Previous studies have focused on pathological events that occur co-incident or after symptom onset and MN degeneration. For many years, research has centered on the MN cell body in the spinal cord and central nervous system (CNS) as the key site of pathogenesis in ALS, but several studies have found that peripheral (PNS) events may initiate the disease in terms of clinical symptoms, and supportive glial cells in the CNS are also involved in disease pathology. Numerous ALS clinical trials have been unsuccessful, perhaps because the treatments are initiated too late in the course of the disease or because the targeted mechanisms are too far down the cascade of events that leads to motor neuron death. Recent studies characterizing disease pathogenesis in mutant SOD1 FALS mice have revised the traditional view of ALS as a disease of the cell body with secondary loss of axons and synapses. There is now a growing consensus in the field that the axon and synapses are the first cellular sites of degeneration. However, there is still controversy over (1) whether axon and synapse loss is initiated at those sites or by pathology in the cell body, in non-neuronal cells or even in non-MNs and (2) the specific molecular mechanisms mediating axon/synapse loss in ALS are largely unknown. To investigate these issues we are proposing the following specific aims: Aim 1: To identify the events that occur before symptom onset in the mutant SOD1G93A mouse PNS. Aim 2: To identify the events that occur before symptom onset in the mutant SOD1G93A mouse CNS. Aim 3: To determine if similar events occur before symptom onset in other models of motoneuron disease. In summary, the major goals of our proposal are to elucidate the specific mechanisms that initiate and mediate axon/synapse loss by describing disease pathogenesis and to identify strategies for preventing axon/synapse loss. In this way we hope to open up new avenues for the treatment of motor neuron disease and other neurodegenerative diseases that involve early loss of axons and synapses. PUBLIC HEALTH RELEVANCE: We are proposing to study early changes in morphology and protein expression at the NMJ and relate these events with prior or subsequent changes at the cell body. The research uses a multidisciplinary approach to address the fundamental question of which portion of the nervous system demonstrates the initial changes of ALS. The results of this study will also be used for patient-oriented research that we are poised to begin. We anticipate that after identifying the location of early nervous system changes in ALS we will use this information to propel further studies into disease biomarkers and the development of therapeutic interventions.